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Caoutchouc tree anther callus vitrification ultra-low temperature storage method

A technology of vitrifying ultra-low temperature and callus, applied in the field of plant cell engineering, can solve problems such as inconvenience of operation, and achieve the effects of convenient operation, simple equipment, good cryopreservation effect and repeatability

Inactive Publication Date: 2009-01-28
RUBBER RES INST CHINESE ACADEMY OF TROPICAL AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

They lowered the temperature of the embryogenic callus and its environment by 0.2°C per minute, and when the temperature dropped to -40°C, they were put into liquid nitrogen for storage, which was inconvenient to operate

Method used

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Examples

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Embodiment Construction

[0030] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

[0031] Various media and required solutions were prepared in advance as required. During the two flowering seasons of spring and summer, rubber trees are in full flowering stage, and the flowers of rubber trees with high yield and good character are selected, and the anthers whose pollen development state is uninucleate stage and a few are binucleate stage are selected. Flower buds are sterilized with 10% bleach solution or 0.1% mercuric chloride solution for 10 minutes, rinsed with sterile water for 4 to 6 times, take out anthers under aseptic conditions, and put them on embryogenic callus induction medium (MS medium Add 2,4-D 1.0mg / L, KT 0.6mg / L, NAA 0.5mg / L, Phytagel 2.5g / L, sucrose 70.0g / L, coconut water 70ml / L.) to induce embryogenic healing damage tissue. The callus was transferred to the pre-medium prepared in advance accordin...

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Abstract

The invention provides a method for preserving the callus of rubber plant anther with vitrification and super low temperature, comprising the following steps: the anther callus is inoculated to the pre-culture medium, dark culture is carried out for 2 to 3 days with the culture temperature of 25 to 28 DEG C; the anther callus is processed by 60 percent of PVS2 for 10 to 30 minutes at room temperature; after dehydration process, the anther callus is processed by the PVS2 for 20 to 40 minutes at the temperature of 0 DEG C, the stock solution of the PVS2 is removed, then new PVS2 is added, and the anther callus is stocked in liquid nitrogen; and after the anther callus is resurgent, successive transfer culture is carried out, the regeneration plant can be obtained after further embryoid is induced and the mature embryoid is induced to carry out differentiation. The preservation method by liquid nitrogen with vitrification and super low temperature related by the invention has the advantages of simple equipment, simple experiment procedure and good effect and repetitiveness, etc., can preserve the callus of rubber plant anther for long time, obtains the regeneration plant by culturing after unfrozen and creates conditions for the genetic transformation of the rubber plant.

Description

technical field [0001] The invention belongs to the technical field of plant cell engineering, and relates to a vitrification ultra-low temperature preservation method of rubber tree anther callus. Background technique [0002] Rubber trees are perennial tall trees with cross-pollination and high genetic heterozygosity, which makes the breeding of rubber trees more difficult. There are generally two kinds of rubber tree planting materials in production, one is the seed tree, and the other is the bud grafting tree. Bud grafting trees have little variation in yield among individuals, high yield and consistent traits, but weak stress resistance. Seedling trees have large yield variation, low yield, and inconsistent traits among individuals, but they have strong resistance to stress. At present, the production of rubber trees is basically done by budding trees. The selection and breeding of rubber trees adopts the method of multi-generation self-crossing according to the conv...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/04
Inventor 李哲周权男孙爱花黄天带黄华孙林位夫
Owner RUBBER RES INST CHINESE ACADEMY OF TROPICAL AGRI SCI
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